Mammalian spermatozoa contain a cholinergic system consisting of choline acetyltransferase (ChA), acetylcholine (ACh), acetylcholinesterase (AChE) and a cholinergic receptor (ChR). They also contain phosphotidylethanolamine (PE) and phosphotidylcholine (PC). PE may be converted to PC by two phosphatide-N-methyltransferases. Choline may be liberated from PC through a sequence of enzymatic steps. This liberated choline may be converted to ACh, which acts on ChR to regulate sperm motility. Thus, ACh synthesis may be linked to phospholipid metabolism. Therefore, the primary objective of this investigation is to demonstrate the operation of an ACh-PC cycle and its components in spermatozoa and to study its significance to sperm motility, viability and fertilizing capacity. The specific aims include: (1) demonstration of two phosphatide-N-methyl-transferases in mammalian spermatozoa, which convert PE to PC, (2) demonstration of the liberation of choline from PC, which will be utilized by spermatozoa to form ACh, (3) evidence for a choline uptake system in spermatozoa, (4) distribution of the various enzymes (ChA, AChE, phospholipases A,B, and D, glycerylphosphorylcholine diesterase, of an ACh-PC cycle in the sperm fractions (heads, midpieces and tails) and seminal plasma; (5) distribution of nicotine and ChA inhibitors (2-benzoylethyltrimethylammonium and related compounds) in sperm fractions and seminal plasma; (6) distribution of enzymes of ACh-PC cycle, nicotine and ChA inhibitors in capacitated sperm; (7) effects of ChA inhibitors, which are tertiary ammonium compounds, in sperm motility in vitro and in vivo; and (8) effects of antifertility agents (cyproterone acetate, alpha-chlorihydrin and trimethylphosphate) on levels of ACh, ChA and acetate thiokinase, ATP citrate lyase, and phosphatide-N-methyltransferases in epididymal spermatozoa in the rat. The results of this investigation may be useful to (1) explain the role of an ACh-PC cycle in the regulation of sperm motility; (2) understand certain types of infertility in men who ejaculate immotile sperm; (3) explain transient infertility among male smokers; and (4) develop antifertility agents.